The problem of drug-resistant bacterial infections has reached epidemic proportions. Methicillin-resistant Staphyolcoccus aureus (MRSA) infections now kill more US citizens each year than does AIDS, and new strategies to treat or prevent such infections are greatly needed. This issue is compounded by a lack of new drug candidates in the pipeline, and the fact that bacteria very quickly develop resistance to new antibiotics. With this project, we propose to explore an alternative strategy for combating bacterial infections, the use of a botanical medicine containing multiple different compounds with diverse modes of action. The botanical under investigation is goldenseal (Hydrastis canadensis), which has been used in traditional and modern alternative medicine to treat infections. Goldenseal is a significant topic of study because of its popularity as a dietary supplement (among the top 15 best-selling botanical medicines in the US, and the 6th most commonly used by children). This plant or its constituents have demonstrated antimicrobial activity in numerous in vitro and several in vivo studies. Our strong preliminary data provide new insights into the mechanisms by which goldenseal acts against bacteria. We have shown that this plant contains a diverse array of bioactive constituents, which can act as direct antimicrobia agents, as efflux pump inhibitors, and as anti-virulence compounds (via quorum quenching). A major goal of this application is to identify the constituents that contribute to these diverse activities. To accomplish this, we will employ an innovative approach, synergy-directed fractionation, which has recently been developed and published by the principal investigator's laboratory. Our long term objective is to use the information provided by these studies to enable the preparation of topical goldenseal preparations with optimal efficacy for the treatment or prevention of bacterial infections. More broadly, we also seek to explore one of the major guiding principles of complementary and alternative medicine, that the activity of multi-component therapies can be enhanced due to synergistic interactions. The methods validated as a part of this study are expected to prove useful to other investigators who must account for the synergistic interactions that play a role in the activity of many botanical medicines.